DNA endonucleases isolated from bacteria. Characterized by ability
to recognize specific sequences on DNA - a
restriction site or restriction sequence - and cut
at exactly the same place every time.
RE always cut sugar-phosphate backbone so as to leave a 5’ phosphate and
a 3’ hydroxyl.
Some RE make staggered cuts leaving a
5’ phosphate extension and a recessed
3’ hydroxyl end. (EcoRI; BamHI).
Some RE making staggered cuts leave a 3’
hydroxyl extension and a recessed
5’ phosphate end. (PstI).
Some RE leave blunt ends (HaeIII;
HpaI)
Restriction sites are palindromic
sequences; they read same 5’ -- 3’ on both strands.
Restriction sites can be 4, 6 or 8 base pairs.
Some restriction sites may be degenerate.
At a certain position the base can be
either of the 2 purines or
either of the 2 pyrimidines.
Sometimes it can be any of the 4 nucleotides.
Number of sites per genome depends on number of bases in recognition sequence
- 1/4 n
For 4 base cutter = 1 site in 256 bases (11,718,750 restriction sites
in a 3000 MB genome).
For 6 base cutter = 1site in 4, 167 bases (719,942 restriction
sites in a 3000 MB genome).
For 8 base cutter = 1 site in every 66,667 bases (45,000 restriction
sites in a 3000 MB genome).
RE ppear to be a mechanism to protect bacteria from infection by bacteriophages
and foreign DNA
Naming convention - first letter genus; second, third letter species; roman
numeral order of discovery.
Some RE cut at different places on each strand and leave staggered cuts.
The single stranded regions form “sticky ends”. This makes them
very useful as a tool for cloning DNA!
Type I and Type III restriction enzymes recognize specific sites, but cut
far outside of these sites. Since the sequence outside of the
recognition sites can be anything, the “sticky ends” are essentially random
and are much less useful.
